CN104480025B - A kind of alternaric bacteria Y1309 1 and its application - Google Patents
A kind of alternaric bacteria Y1309 1 and its application Download PDFInfo
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- 239000002245 particle Substances 0.000 claims abstract description 19
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- 238000000034 method Methods 0.000 claims abstract description 10
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- 238000006243 chemical reaction Methods 0.000 abstract description 8
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- 239000000126 substance Substances 0.000 description 5
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- 238000013461 design Methods 0.000 description 4
- 239000007788 liquid Substances 0.000 description 4
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 4
- 238000002360 preparation method Methods 0.000 description 4
- 230000002829 reductive effect Effects 0.000 description 4
- 229910052709 silver Inorganic materials 0.000 description 4
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- 229910052793 cadmium Inorganic materials 0.000 description 2
- BDOSMKKIYDKNTQ-UHFFFAOYSA-N cadmium atom Chemical compound [Cd] BDOSMKKIYDKNTQ-UHFFFAOYSA-N 0.000 description 2
- 239000008367 deionised water Substances 0.000 description 2
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- 238000004519 manufacturing process Methods 0.000 description 2
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- 229910052697 platinum Inorganic materials 0.000 description 2
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- 108010024636 Glutathione Proteins 0.000 description 1
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 1
- 239000001888 Peptone Substances 0.000 description 1
- 108010080698 Peptones Proteins 0.000 description 1
- 241000191940 Staphylococcus Species 0.000 description 1
- 235000009754 Vitis X bourquina Nutrition 0.000 description 1
- 235000012333 Vitis X labruscana Nutrition 0.000 description 1
- 240000006365 Vitis vinifera Species 0.000 description 1
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- 238000011143 downstream manufacturing Methods 0.000 description 1
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- 230000002538 fungal effect Effects 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 229960003180 glutathione Drugs 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 229910052737 gold Inorganic materials 0.000 description 1
- 239000010931 gold Substances 0.000 description 1
- 230000005764 inhibitory process Effects 0.000 description 1
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- 239000002105 nanoparticle Substances 0.000 description 1
- 235000019319 peptone Nutrition 0.000 description 1
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- 108090000623 proteins and genes Proteins 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
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- 230000006641 stabilisation Effects 0.000 description 1
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- 238000001308 synthesis method Methods 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 238000010189 synthetic method Methods 0.000 description 1
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- C12N1/145—Fungal isolates
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Abstract
The invention discloses a kind of alternaric bacteria Y1309 1, it prepares the method mild condition of Nano Silver, without using a variety of chemical reagent, and reaction cost is low, environment-friendly.The invention also discloses applications of the alternaric bacteria Y1309 1 in terms of metal nano material is prepared.The invention also discloses applications of the alternaric bacteria Y1309 1 in terms of Nano Silver is prepared.Nano-Ag particles uniformity produced by the present invention is good, can independently disperse in water, stable under normal temperature.The Nano Silver that the present invention is produced has certain inhibitory activity to Escherichia coli and silver-colored staphylococcus aureus.The present invention can be applied in fields such as medicine, catalysis, environment remediations.
Description
Technical field
The present invention relates to biological technical field, and in particular to a kind of alternaric bacteria Y1309-1 and its application.
Background technology
Nano Silver has the characteristics such as unique heat, light, electricity, magnetic, catalysis and sensitivity, catalyst material, antistatic material,
It is widely used in terms of low temperature superconducting material, electric slurry and biology sensor material, biomedicine.
The synthetic method of current Nano Silver has Physical, chemical method and bioanalysis.Physical requires high to instrument and equipment, raw
Somewhat expensive is produced, and power consumption is high.The chemicals that chemical method is used can be polluted to environment, can also be adsorbed on the surface of Nano Silver
And it is influenceed in the application of field of medicaments；And its production Nano Silver can be over time extension assemble, therefore storage meeting
Influence the particle diameter of Nano Silver.Biological synthesis method is because of gentle, environment-friendly, the nanometer of synthesis with low cost, reaction condition
Particle size distribution is narrow, stability is high, good biocompatibility the advantages of, it has also become an important branch of nanometer synthetic technology.
Microorganism has the advantages that NATURAL DISTRIBUTION is wide, be separately cultured simple, growth metabolism is fast, strong adaptability, easily operated,
It is the green bio factory of nano materials.And fungi is larger because its hyphal surface is accumulated, substantial amounts of extracellular protein can be secreted
Matter, is favorably improved Nano Silver yield；And its production process is economic and environment-friendly, economic feasibility is good, and Downstream processing is convenient, receives
More concerns.
The content of the invention
Goal of the invention：In view of the deficienciess of the prior art, first purpose of the present invention is to provide a kind of alternaric bacteria
Y1309-1, it prepares the method mild condition of Nano Silver, without using a variety of chemical reagent, and reaction cost is low, environment-friendly.
Second purpose of the invention there is provided applications of the alternaric bacteria Y1309-1 in terms of metal nano material is prepared.The present invention the
Three purposes there is provided applications of the alternaric bacteria Y1309-1 in terms of Nano Silver is prepared.Nano-Ag particles produced by the present invention
Uniformity is good, can independently disperse in water, stable under normal temperature.The Nano Silver that the present invention is produced is to Escherichia coli and silver yellow color Portugal
Grape coccus has certain inhibitory activity.The present invention can be applied in fields such as medicine, catalysis, environment remediations.
Technical scheme：To realize the purpose of foregoing invention, the technical solution adopted by the present invention is：A kind of alternaric bacteria
Y1309-1, its Classification And Nomenclature is alternaric bacteria Y1309-1（Alternaria sp. Y1309-1）, the bacterial strain is in 2014 7
It is preserved in China typical culture collection center the moon 3, deposit number is CCTCC No：M 2014311.Preservation address：Hubei
Wuhan University of wuchang, wuhan area of province collection (the first affiliated primary school of Wuhan University opposite).
The alternaric bacteria bacterium colony is just white, fades to taupe, mycelia is sparse, matrix dark brown.Single of conidiophore,
Different in size, basidixed not branch, conidium has a barrier film in length and breadth, the shape of falling club, avette.
Above-mentioned alternaric bacteria Y1309-1 screening technique, it is characterised in that comprise the following steps：
1）35 fungal strains for being isolated from Southeast China University's campus soil of Laboratories Accession are taken, respectively in PDA solid mediums
On in 28 DEG C cultivate 3-5 days；
2）35 plants of strains are inoculated in 100mL potato dextrose broths respectively, in 28 DEG C, 150rpm cultures 2
It obtains thalline；
3）Collect thalline and use sterile water washing, 20g wet thallus is added in 100ml aseptic deionized waters, in 28 DEG C, pH
7th, 150rpm cultivates 48h；
4）It is centrifuged off step 3）In thalline, collect supernatant, final concentration 0.5-2.0 mmol/L silver nitrate is molten
Liquid adds step 4）Supernatant in, in 28 DEG C, 150rpm reaction 12-48h, according to the color of solution after reaction judge whether life
Into Nano Silver（It is yellowish-brown to dark brown to generate the solution of Nano Silver）, screening experiment is repeated and to life to the system reacted
Into Nano Silver confirmed；
5）Collection step 3）In thalline, after sterile water washing, 20g wet thallus is added into 100ml final concentrations 0.5-2.0
In mmol/L sterile silver nitrate solution, 12-48h is reacted in 28 DEG C, pH 7,150rpm, is sentenced according to the color of thalline after reaction
Whether its intracellular of breaking generates Nano Silver（It is yellowish-brown to dark brown to generate the cell of Nano Silver）, the system reacted is repeated
Screening experiment and Nano Silver to generation is confirmed；Obtained one plant of guarantor of the bacterial strain that can generate Nano Silver selection will be screened
Hide, and be named as alternaric bacteria Y1309-1.
Applications of the above-mentioned alternaric bacteria Y1309-1 in terms of metal nano material is prepared.
Applications of the above-mentioned alternaric bacteria Y1309-1 in terms of Nano Silver is prepared.
Above-mentioned application, the extracellular method and steps for preparing Nano Silver of alternaric bacteria Y1309-1 are as follows：
1）Alternaric bacteria Y1309-1 is cultivated 4 days on PDA solid mediums in 28 DEG C；
2）Alternaric bacteria Y1309-1 is inoculated in 100mL potato dextrose broths, trained in 28 DEG C, 150rpm
Support and obtain thalline in 2 days；
3）Collect thalline and use sterile water washing, 20g wet thallus is added in 100ml aseptic deionized waters, in 28 DEG C, pH
7th, 150rpm cultivates 48h；
4）It is centrifuged off step 3）In thalline, collect supernatant；
5）Final concentration 0.5-2.0 mmol/L silver nitrate solution is added into step 4）Supernatant in, in 28 DEG C,
150rpm reacts 12-48h, obtains nano-Ag particles, and its particle diameter is 40-50nm.
Above-mentioned application, the method and step that the alternaric bacteria Y1309-1 intracellulars prepare Nano Silver is as follows：
1）Alternaric bacteria Y1309-1 is cultivated 4 days on PDA solid mediums in 28 DEG C；
2）Alternaric bacteria Y1309-1 is inoculated in 100mL potato dextrose broths, trained in 28 DEG C, 150rpm
Support 2 days；
3）Thalline is collected, after sterile water washing, 20g wet thallus is added 100ml final concentration 0.5-2.0 mmol/L's
In sterile silver nitrate solution, 6-48h is reacted in 28 DEG C, pH 7,150rpm, Nano Silver is made in intracellular；
4）Broken thalline, collects nano-Ag particles.
Generally believe that the bioactive substance that microorganism produces includes protein, reduced sugar, reductive glutathione at present
Nano material is reduced into etc. that metal ion such as silver, cadmium, palladium, platinum can be carried out into enrichment, or by two metal ion species in system
Nanoalloy is reduced into, and biomolecule plays an important role to nano metal stabilization.Therefore, the alternaric bacteria in the present invention
There can be the ability that gold, cadmium, palladium, platinum plasma reduction are assembled into nano-particle or alloy.
Beneficial effect：There is advantages below compared with prior art：Nano-Ag particles uniformity produced by the present invention is good, can
It is stable under normal temperature independently to disperse in water.The Nano Silver that the present invention is produced has to Escherichia coli and silver-colored staphylococcus aureus
Certain inhibitory activity.The present invention can be applied in fields such as medicine, catalysis, environment remediations.
Brief description of the drawings
Fig. 1 is the extracellular design sketch for preparing Nano Silver of alternaric bacteria Y1309-1 of embodiment 2；
Fig. 2 prepares the design sketch of Nano Silver for the alternaric bacteria Y1309-1 intracellulars of embodiment 3；
Fig. 3 is the UV-vis spectroscopy spectrogram of the Nano Silver of the extracellular preparations of alternaric bacteria Y1309-1 of embodiment 2；
Fig. 4 is the lens TEM photos of the silver nano-grain of the extracellular preparations of alternaric bacteria Y1309-1 of embodiment 2；
Fig. 5 is the lens SEM photograph of the silver nano-grain of the extracellular preparations of alternaric bacteria Y1309-1 of embodiment 2；
Fig. 6 prepares the lens TEM photos of Nano Silver for the alternaric bacteria Y1309-1 intracellulars of embodiment 3；
Fig. 7 is the design sketch that Nano Silver prepared by embodiment 2 suppresses Escherichia coli；
Fig. 8 is the design sketch that Nano Silver prepared by embodiment 2 suppresses golden yellow staphylococcus.
Embodiment
Present disclosure is further illustrated below in conjunction with accompanying drawing and specific implementation, but should not be construed as the limit to the present invention
System.Without departing from the spirit and substance of the case in the present invention, the modifications or substitutions made to the inventive method, step or condition,
Belong to the scope of the present invention.
Embodiment 1：Alternaric bacteria Y1309-1 condition of culture
Alternaric bacteria Y1309-1 is in PDA solid mediums（Potato 20%, glucose 2%, deionized water 1000mL, pH are certainly
So, 121 DEG C of sterilizing 30min are stand-by）On in 28 DEG C culture, liquid medium within（PDA solid mediums：Potato 20%, grape
Sugar 2%, agar 1.5%, deionized water 1000mL, pH are naturally, 121 DEG C of sterilizing 30min are stand-by）In in 28 DEG C, 150rpm cultivate.
The alternaric bacteria Y1309-1 of embodiment 2 is extracellular to prepare Nano Silver
Step 1 cultivates alternaric bacteria 4 days on PDA solid mediums in 28 DEG C；
Alternaric bacteria is inoculated in 100mL potato dextrose broths by step 2, in 28 DEG C, 150rpm cultures 2
My god；
Step 3 collects thalline and uses sterile water washing, 20g wet thallus is added in 100ml aseptic deionized waters, in 28
DEG C, pH 7,150rpm culture 48h；
Step 4 removes thalline, collects supernatant；
Step 5 is by silver nitrate solution（Final concentration 0.5-2.0 mmol/L）Add in above-mentioned supernatant, in 28 DEG C, 150rpm
12-48h is reacted, nano-Ag particles are obtained, its particle diameter is 40-50nm.
The left supernatant to add before silver nitrate of accompanying drawing 1, the right side is molten to add the dark brown Nano Silver after nitric acid silver reaction 48h
Liquid.Accompanying drawing 3 is the UV-vis spectroscopy spectrogram of Nano Silver prepared by embodiment 1.Accompanying drawing 4 and accompanying drawing 5 are respectively alternaric bacteria
The ESEM and transmission electron microscope photo of the nano-Ag particles of extracellular preparation, display dispersed nano-silver particles are good, and particle is equal
Even, size is generally 40-50nm or so.
The microorganism alternaric bacteria Y1309-1 intracellulars of embodiment 3 prepare Nano Silver
Step 1 cultivates alternaric bacteria 4 days on PDA solid mediums in 28 DEG C；
Alternaric bacteria is inoculated in 100mL potato dextrose broths by step 2, in 28 DEG C, 150rpm cultures 2
My god；
Step 3 collects thalline, after sterile water washing, and 20g wet thallus is added into the sterile silver nitrate solutiones of 100ml（It is dense eventually
Spend 0.5-2.0 mmol/L）In, 6-48h is reacted in 28 DEG C, pH 7,150rpm, Nano Silver is made in intracellular；
Step 4 crushes thalline, collects nano-Ag particles.
The left thalline to add before silver nitrate of accompanying drawing 2, the right side is the bacterium of the parcel Nano Silver added after nitric acid silver reaction 48h
Body, is yellowish-brown.
Accompanying drawing 6 is the transmission electron microscope photo of nano-Ag particles prepared by alternaric bacteria intracellular, and display nano-Ag particles are uniform.
The bacteriostasis of Nano Silver synthesized by the embodiment 2 of embodiment 4 to bacterium
Escherichia coli and staphylococcus aureus are coated on solid medium by step 1（Beef extract 0.5%, NaCl
0.5%, peptone 1%, pH 7.6 ~ 7.8）；
Step 2 places a sterilizing filter paper on solid medium, and the Nano Silver added thereon prepared by embodiment 2 is molten
The μ l of liquid 5；
Escherichia coli and staphylococcus aureus are cultivated 24h by step 3 in 37 DEG C, and its antibacterial circle diameter is measured respectively and is taken
Average value, determines that antibacterial circle diameter of the Escherichia coli under Nano Silver effect is 1.51cm, staphylococcus aureus is in Nano Silver
Antibacterial circle diameter under effect is 1.40cm.
Accompanying drawing 7 and accompanying drawing 8 are that the Nano Silver synthesized by embodiment 2 is big to Escherichia coli and silver-colored staphylococcus aureus inhibition zone
It is small, illustrate that Nano Silver prepared by alternaric bacteria Y1309-1 all has preferably suppression to gram-positive bacteria and Gram-negative bacteria
Make and use.
Claims (4)
1. a kind of alternaric bacteria Y1309-1, it is characterised in that its Classification And Nomenclature is alternaric bacteria Y1309-1（Alternaria sp. Y1309-1）, the bacterial strain is preserved in China typical culture collection center on July 3rd, 2014, and deposit number is
CCTCC No：M 2014311.
2. applications of the alternaric bacteria Y1309-1 in terms of Nano Silver is prepared described in claim 1.
3. application according to claim 2, it is characterised in that the alternaric bacteria Y1309-1 is extracellular to prepare Nano Silver
Method and step is as follows：
1）Alternaric bacteria Y1309-1 is cultivated 4 days on PDA solid mediums in 28 DEG C；
2）Alternaric bacteria Y1309-1 is inoculated in 100mL potato dextrose broths, cultivated 2 days in 28 DEG C, 150rpm
Obtain thalline；
3）Collect thalline and simultaneously use sterile water washing, by 20g wet thallus addition 100ml aseptic deionized waters, in 28 DEG C, pH 7,
150rpm cultivates 48h；
4）It is centrifuged off step 3）In thalline, collect supernatant；
5）Final concentration 0.5-2.0 mmol/L silver nitrate solution is added into step 4）Supernatant in, in 28 DEG C, 150rpm it is anti-
12-48h is answered, nano-Ag particles are obtained, its particle diameter is 20-60nm.
4. application according to claim 2, it is characterised in that the alternaric bacteria Y1309-1 intracellulars prepare Nano Silver
Method and step is as follows：
1）Alternaric bacteria Y1309-1 is cultivated 2-4 days on PDA solid mediums in 28 DEG C；
2）Alternaric bacteria Y1309-1 is inoculated in 100mL potato dextrose broths, in 28 DEG C, 150rpm cultures 2
My god；
3）Thalline is collected, after sterile water washing, 20g wet thallus is added into the sterile of 100ml final concentration 0.5-2.0 mmol/L
In silver nitrate solution, 6-48h is reacted in 28 DEG C, pH 7,150rpm, Nano Silver is made in intracellular；
4）Broken thalline, collects nano-Ag particles.
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